Induction of cyclic AMP synthesis by forskolin is followed by a reduction in the expression of c‐myc messenger RNA and inhibition of 3H‐thymidine incorporation in … (original) (raw)
Related papers
Journal of Cellular Physiology, 1989
We have studied the effect of increased intracellular levels of cyclic AMP on the growth response to platelet-derived growth factor (PDGF) of human foreskin fibroblasts in culture. It was found that forskolin, a potent stimulator of adenylate cyclase activity, inhibits the stimulatory effect of PDGF on 3H-thymidine incorporation with a dose dependence similar to that observed with regard to cyclic AMP formation. A time-course study indicated that forskolin has no effect on ongoing DNA synthesis but affects events in the prereplicative phase. The cell-cycle block induced by forskolin was found to be reversible; after removal of the drug, DNA synthesis was initiated after a lag period, similar to that of the prereplicative phase of control cells. Forskolin had no effect on PDGF binding, receptor autophosphorylation, or c-fos mRNA expression. However, a reduction in PDGF-induced c-myc mRNA expression was observed in cultures given forskolin. Forskolin was also found to have a marked stimulatory effect on the expression of interferon-p2 mRNA expression. However, we were unable to demonstrate that the growth-inhibitory effect of forskolin i s mediated by interferon+. In conclusion, an increase in CAMP levels leads to a reversible inhibition of PDGF-induced DNA synthesis in human fibroblasts, which may be related to an inhibition of c-myc mRNA expression. Cells were cultured in Eagle's minimum essential medium, supplemented with 10% newborn calf serum (GIBCO) and antibiotics (100 U penicillin and 50 pg streptomycin per ml) in a humidified atmosphere containing 5% COz. For experiments, subconfluent cell cultures were serum-starved for 2-4 days in MCDB 104 medium containing 0.5 mM Ca2+.
A cAMP independent inhibitory action of high doses of forskolin in rat Leydig cells
The Journal of Steroid Biochemistry and Molecular Biology, 1990
In addition to well known direct stimulatory and potentiatory actions of forskolin, we have previously reported that low doses of this diterpene (10-9, 10-12 M) markedly inhibit the production of cAMP and testosterone in rat Leydig cells through a pertussis toxin sensitive G-protein (A. Khanum and M. L. Dufau, J. Biol. Chem. 261, 1986). A different type of inhibitory effect of forskolin is described in this study. Forskolin (10-5 M) markedly stimulates basal adenylate cyclase activity (about 200%) in rat Leydig cell membranes and potentiates the stimulatory effect of gonadotropin (10-9, 10-7 M) on adenylate cyclase in presence or in absence of GTP (10-5 M). Similarly a time-dependent stimulation of forskolin (10-~ M) alone is noted on all cAMP pools and testosterone production. Using a supramaximal steroidogenic dose of hCG (0.26 nM) or choleragen (0.1/~M), forskolin potentiates the gonadotrophin and toxin-induced responses of all cAMP pools significantly while inhibiting testosterone production. Moreover, forskolin also inhibits 8-Bromo-cAMP stimulated steroidogenesis. In contrast, pregnenolone synthesis was not altered by the diterpene. We have demonstrated in this study that the inhibitory effect of high doses of forskolin on steroidogenesis is distal to cAMP generation, and resulted from a steroidogenic block residing beyond pregnenolone synthesis.
Prostaglandins, 1986
Recent experimental studies indicated that prostaglandin E 2 (PGE 2) is the most abundant prostanoid synthesized by rabbit articular chondrocytes. Exogenous PGE 2 stimulates cyclic AMP (cAMP) synthesis in these cells. Analogues of cAMP and forskolin have now been shown to suppress the biosynthesis of PGE 2 in the presence of serum in a time-dependent manner. The most abundant prostanoid, PGE 2 was most markedly affected. PGF2~ was unaffected. These results indicated that intracellular accumulation of cAMP in chondrocytes and relative resistance of cAMP to phosphodiesterases control prostanoid synthesis in a negative feedback loop.
Forskolin induces U937 cell line differentiation as a result of a sustained cAMP elevation
European Journal of Pharmacology, 1998
. The present study examines the effects of forskolin on U937 cell differentiation. We recently reported that dibutyryl cAMP dbcAMP , but not cAMP-elevating agents such as histamine, promotes U937 cell differentiation. cAMP production elicited by stimulation of histamine H receptors showed a rapid, homologous desensitization, which might explain the dissimilar responses to histamine and 2 Ž . dbcAMP. Forskolin induced an increase in cAMP levels in a concentration-dependent manner EC s 30 mM for an extended period of 50 Ž . Ž . at least 24 h. Forskolin but not histamine up to 100 mM , also inhibited cell growth in a dose-dependent fashion EC s 22 mM . After 50 3 days of incubation, 75 mM forskolin induced U937 cell differentiation as judged by an increased rate of reduction of nitrobluetetra-Ž . zolium mean " S.E.M.: 21.3 " 6.6% in treated cells vs. 3.2 " 1.9% in the control group, P -0.001 and an augmented chemotactic Ž . Ž . response to complement 5a C5a 33.2 " 5.9% in forskolin-treated vs. 0.34 " 0.12% in control cells, P -0.01 . Furthermore, c-Myc levels decreased following forskolin treatment, while the histamine H receptor agonist dimaprit had no effect. We conclude that 2 forskolin induces U937 cell differentiation through a sustained rise in cAMP levels. q
DNA synthesis in cultured human fibroblasts: Regulation by 3′ : 5′-cyclic amp
Journal of Supramolecular Structure, 1976
The addition of serum t o density-inhibited human fibroblast cultures induced a wave of DNA synthesis, measured as [ HI thymidine incorporation into acidprecipitable material, beginning after !-12 hr and reaching maximum levels at 16-24 hr. Addition of dibutyryl-3': 5-cyclic AMP(DBcAMP) together with serum inhibited ['HI thymidine incorporation by 75-95%. When DBcAMP was added for the first 4 hr of serum stimulation and then removed, the wave of DNA synthesis was not delayed. This suggested that serum could induce DNA synthesis even though cyclic AMP concentrations were maintained at high levels by DBcAMP during this initial period. These results ar: inconsistent with the hypothesis that it is the immediate transient reduction in 3': 5-cyclic AMP concentration following the addition of serum that triggers DNA synthesis. By contrast, DBcAMP added 8 hr after serum inhibited ['HI thymidine incorporation t o the same extent as DBcAMP added at the same time as serum. This indicated that a step essential for DNA synthesis and occurring late in G I was inhibited by high concentrations of 3' : 5'-cyclic AMP. I NTRO DUCT1 ON Density-inhibited human fibroblast cultures can be induced t o synthesize DNA by growth-promoting agents such as fetal calf serum, insulin, and certain polypeptides with insulin-like activity (1). The stimulation by serum appeared to proceed by a mechanism different from that by insulin or insulin-like polypeptides (1). Addition of insulin or serum 199 (159)
Biomedicine & Pharmacotherapy, 1992
Eight-bromo cyclic AMP (8-Br-CAMP) and substances elevating CAMP levels within cells such as foskolin, cholera toxin and Brodetellu pertussis invasive adenylate cyclase (BPAC) suppress growth of cultured granulosa cells co-transfected by SV40 DNA and Ha-ras oncogene concomitantly with the induction of steroidogenesis and without affecting the oncogene expression [l-31. We therefore tested the hypothesis that CAMP can modulate tumorigenesis and metastatic spread of these cells in vivo. The co-transfected cells induced rapid development of tumors when injected subcutaneously in nude mice. Tumor development was faster in less differentiated co-transfected cells originating from preantraI ovarian foBides than in those obtained from highly differentiated transformed cells originating from preovulatory follicles. Cells transfected by SV40 DNA alone produced only slow growing small tumors. Metastatic lesions of co-transfected cells were most abundant in the lung and less frequent in ovaries, kidney and spleen. No metastatic lesions were found in the liver. However, metastatic spread was dramatically suppressed when co-transfected cells injected into nude mice were pretreated with the invasive BPAC. In contrast, no suppression of metastases was observed when the cells were pretreated with 8-Br-CAMP, forskolin or cholera toxin. Removal of forskolin in cultured co-transfected cells yielded a rapid decrease in CAMP levels. In contrast, high levels of CAMP persist in cell cultures even several hours after 1 h pretreatment and subsequent removal of BPAC from the medium of cultured co-transfected cells. It is suggested that the inhibitory effect of BPAC on metastatic spread of these cells is due to prolonged elevation of CAMP in vivo. The newly established granulosa cell lines transformed by SV40 and the Ha-rus oncogene can serve as a model for further studies on CAMP modulation of carcinogenesis in ovarian malignancies.